Mitogen-activated protein kinase (MAPKs) (also called extracellular signal-regulated kinases or ERKs) are rapidly activated in response to ligand bindings by both growth factor receptors that are tyrosine kinases (such as the epidermal growth factor (EGF) receptor) and receptors that are coupled to heterotrimeric guanine nucleotide binding proteins (G proteins) such as the thrombin receptor. In addition, receptors like the T cell receptor (TCR) and B cell receptor (BCR) are non-covalently associated with src family tyrosine kinases which activate MAPK pathways. Specific cytokines such as tumor necrosis factor (TNFα) can also regulate MAPK pathways. The MAPKs appear to integrate multiple intracellular signals transmitted by various second messengers. MAPKs phosphorylate and regulate the activity of enzymes and transcription factors including the EGF receptors Rsk 90, phospholipase A2, c-Myc, c-Jun and Elk-1/TCF. Although the rapid activation of MAPKs by receptors that are tyrosine kinases is dependent on Ras, G protein-mediated activation of MAPK appears to occur through pathways dependent and independent of Ras.
The MAPKs are activated by phosphorylation on both a threonine and tyrosine by dual specificity kinases. MAPK/ERK kinases (MEKs) which are, in turn, activated by serine/threonine phosphorylation MAPK kinase kinases (MKKKs or MEKKs). At present, at least four MEKKs have been identified. The four MEKK proteins range from 69.5-185 kDa in size, having their kinase domains in the carboxy-terminal end of the protein and their catalytic domains in the amino-terminal end of the protein. Murine MEKK1 was cloned initially on the basis of its homology with the STE11 and Byr2 kinases from yeast (Lange-Carter et al. (1993) Science 260:315-319: Xu et al. (1996) Proc. Natl. Acad. Sci. USA 93:5291-5295; and Blank et al. (1996) J. Biol. Chem. 271:5361-5368). Murine MEKK2 and MEKK3 were subsequently cloned and found to have 94% homology in their kinase domains as well as 65% homology within their catalytic domains. Blank et al., supra. The cloning of murine MEKK4 revealed approximately 55% homology to the kinase domains of MEKKs 1, 2, and 3 whereas the amino-terminal region of MEKK4 has little sequence homology to the other MEKK family members. Gerwin et al. (1997) J. Biol. Chem. 272:8288-8295. MEKK1 and MEKK4, but not MEKK2 and MEKK3, bind to the low molecular weight GTP-binding proteins Cdc42 and Rac. Furthermore, MEKK1 also binds to Ras in a GTP-dependent manner (Russell et al. (1996) J. Biol. Chem. 11757-11760) and Ras activity is required for EGF-mediated stimulation of MEKK1 activity (Lange-Carter and Johnson (1994) Science 265:1458-1461). In addition to growth factor receptor tyrosine kinases (i.e. EGF receptor), the TNF receptor, the FcεR1 in mast cells Ishizuka et al. et al. (1996) J. Biol. Chem. 271:12762-12766) and the N-formyl methionyl leucine peptide receptor in neutrophils have been shown to activate MEKK1. EGF and TNF also activate MEKK3 and it also appears that the other MEKK proteins are regulated by tyrosine kinase receptors but the intermediate components and effector molecules leading to their activation are poorly understood.
The cellular effects of MEKK1 are quite diverse and can be classified as being either JNK-dependent or JNK-independent. For example, MEKK1 can mediate activation of ERK1 and ERK2 and, by a yet undefined mechanism, activation of the c-Myc transcription factor independent of JNK activity (Lassignal-Johnson et al. (1996) J. Biol Chem. 271:3229-3237 and Lange-Carter et al. (1993) Science 260:315-319). Alternatively, MEKK1 may or may not require JNK activity for activation of IκB kinase which leads to NKκB activation (Liu et al. (1996) Cell 87:565-576 and Meyer et al. (1996) J. Biol. Chem. 271:8971-8976). Furthermore, depending upon the cell type, MEKK1, but not MEKK2, 3 or 4, has been shown to mediate apoptosis by both JNK-dependent and JNK-independent mechanisms (Xia et al. (1995) Science 270:1326-1331 and Lassignal-Johnson et al. (1996) J. Biol Chem. 271:3229-3237).
Given the important role of members of the MAPK signal transduction cascade, in particular the MEKK signal transduction molecules, in regulating mammalian cellular processes ranging from cellular proliferation and differation to cellular apoptosis, there exists a need for identifying human MEKK nucleic acid and protein molecules as well as for modulators of such molecules for use in regulating a variety of human cellular responses.